Individual genetic mutations can predispose a cell or tissue toward certain diseases, such as most cancers, cystic fibrosis, and sickle cell anemia. Specific mutations in such genes can thus be used as diagnostic indicators for the susceptibility of disease, aiding in early detection and treatment. The high-throughput detection of genes has been studied for several years and devices like microarrays and the DNA chip have significantly increased our capabilities.
The basic principle of microacrray technology requires tagging of the sample with fluorescent dyes. However, these modifications can change the thermodynamic properties of the molecular interactions of DNA and, in some cases, unnaturally stabilize or destabilize the DNA double-strand and change the melting temperature significantly. Additionally, expensive fluorescent microscopes are needed to visualize the data and normalization of the data to references remains problematic. Further, the hybridization of the probe and target molecules is a diffusion-limited process requiring long-incubation times as the target molecules must travel to the arrayed probes on the surface of the chip. The fluorophores are also known to have great effect on the stability of the duplexes as a function of the sequence itself. In addition, fluorescent dyes photobleach, quench statically, or interact with each other, so the microarray technologists need to have very detailed knowledge about the limitations of the optics, reagents used, and the sample interactions.
Several silicon-based approaches have been reported for chemical and biological sensing. While many of the silicon-based sensors can be fabricated with compact size, none of these efforts have resulted in a portable sensor with adequate performance. The hybridization of DNA with probe-functionalized chip surfaces has been studied for biophysical characterization and kinetics studies but its use in a functional nano-scale device has not been reported. The challenges range from the costly and lengthy fabrication processes to the need for external expensive measurement equipment. The DNA hybridization detection techniques have been implemented on chips but throughput, cost, and multiplexing have not been adequately addressed.
The solid-state DNA interaction detection techniques have also required tagging of target with electrical markers, resulting in possibly altered interactions while requiring sample preparation. The DNA electrical detection techniques mostly employ electrochemical impedance spectroscopy, capillary electrophoresis, or charge perturbation detection using labeled target or labeled probe molecules, surface attached antibodies, and/or with pre-/post-PCR. In label-free detection schemes, the probe DNA is immobilized on the electrodes and impedance is measured in conjunction with or without a reference electrode.
Thus, there is a need for biosensors that overcome these problems. The invention is directed to these, as well as other, important ends.